Biochemistry - The Chemical Reactions of Living Cells, Volume 2 - D. Metzler 1980

How electrons meet oxygen, how ATP is generated in the process, and related phenomena
Electron transport chain and oxidative phosphorylation
Sequence of electron carrier function

In what sequence do the carriers operate through which the electron flow passes? In addressing this question, we will take the scheme presented in Figs. 10–11 as our baseline, although certain details remain unknown. The proposed scheme is based on results obtained using several distinct approaches. First, there is strong evidence that the carriers are arranged in order of increasing reduction potential—from left to right in Figs. 10–11. However, this rule does not necessarily hold strictly true, especially considering that the reduction potentials of carriers in Cell/35.html">Mitochondria may differ from those of isolated Enzymes.

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FIG. 10–11. Scheme illustrating current views on the Electron Transport Chain in mitochondria.

Some known respiratory inhibitors^a

^a For sites of inhibition, see Figs. 10–11.

The dual-wavelength spectrophotometer designed by Chance makes it easy to monitor the oxidation or reduction state of a given carrier in mitochondria [68]. This technique, combined with The Use of specific inhibitors (some of which are indicated in Figs. 10–11 and Table 10-2), has helped establish several sequences within The electron transport chain. For instance, blocking with rotenone or amobarbital prevents the reduction of the cytochrome system by NADH, but does not disrupt reduction by succinate or other substrates that have their own flavoprotein intermediaries. Another approach to elucidating The sequence of carriers involves the use of artificial electron acceptors, some of which are listed in Table 10-3. These bypass various segments of the chain, as shown in Figs. 10–11.

An important method for studying the electron transport chain is the fractionation of mitochondrial membranes into fragments that retain The ability to catalyze individual Reactions of the chain. Numerous Methods are used to prepare submitochondrial particles. The well-known Keilin–Hartri Heart Muscle preparation is obtained by homogenizing mitochondria and precipitating fractions at low pH values. Although the resulting particles have a low cytochrome c content and are incapable of Oxidative Phosphorylation, they actively respire. Using ultrasonication, another type of electron carrier preparation has been obtained. Under an Electron microscope, these particles appear as small membrane-bound vesicles resembling mitochondrial cristae.

Although many detergents exert a strongly denaturing effect on Proteins, certain detergents disrupt mitochondrial membranes while preserving enzymatic activity. Digitonin (Fig. 12-18) is the preferred choice among them, as it selectively disrupts the outer membrane. The remaining inner-membrane fragments retain the capacity for oxidative phosphorylation. Subsequent fractionation of submitochondrial particles is carried out via chemical Treatment. One such Procedure yields a set of "complexes" that catalyze reactions in four different segments of the electron transport chain [69]. The Reactions Catalyzed by complexes I, II, III, and IV are specified in equation (10-10).

Chemical Analysis of the complexes revealed the localization of certain components within the intact chain. For example, complexes I and II were found to have a high iron content, whereas copper was detected in complex IV.

It is now appropriate to discuss the specific role that ubiquinone plays in the electron transport chain. A wealth of evidence indicates that it is a true and essential electron carrier. For instance, E. coli mutants unable to synthesize ubiquinone can grow by fermenting glucose, but fail to survive on substrates such as succinate, the utilization of which requires oxidative processes [70]. Lipid-soluble ubiquinone is present in bacterial and mitochondrial membranes in relatively large amounts (Table 10-1) compared to other electron carriers. It appears to function at the junction of the NADH and succinate Branches of the chain and is generally regarded as a "mobile" carrier, in contrast to "fixed" carriers such as Flavoproteins and Cytochromes. In this respect, ubiquinone plays a role similar to that of NAD⁺, which transfers electrons from various soluble dehydrogenases dissolved in the aqueous matrix to the membrane-bound flavoprotein, NADH dehydrogenase. A third mitochondrial component that may function in a similar manner is cytochrome c. Unlike other cytochromes, it is Water-soluble and easily washed out of mitochondrial membranes. Nevertheless, its ratio to fixed cytochromes is typically 1:1. It is unlikely to diffuse as freely as ubiquinone and NAD⁺.